Traditionally, the most common method of preserving biological substances which are unstable in solution at ambient temperatures, such as proteins and DNA, has been freeze-drying. This process involves placing the substance in solution, freezing the solution, and exposing the frozen solid to a vacuum under conditions where it remains solid and the water and any other volatile components are removed by sublimation. The resulting dried formulation contains the biological substance and any salts or other non-volatile materials added to the solution before drying. This drying method, conventionally used in the absence of effective alternatives, often results in significant losses. Pikal (1994) ACS Symposium 567:120-133. Furthermore, many of the various parameters within the freeze-drying process remain poorly characterized, sometimes resulting in the loss of whole batches at the production level.
In spite of the apparent ubiquity of freeze-drying, many freeze-dried substances are still unstable at ambient temperatures. Pikal (1994); Carpenter et al. (1994) ACS Symposium 567:134-147. Damage caused by this process may be circumvented, to a certain degree, by the use of cryoprotectants. Carpenter et al. (1994). However, cryoprotectants may subsequently react with the dried substance. This imposes inherent instability upon storage of the freeze-dried substances.
Other methods used to prepare dry, stable preparations of labile biological and chemical substances such as ambient temperature drying, crystallisation or co-precipitation also have drawbacks. Ambient temperature drying techniques eliminate the freezing step and associated freeze-damage to the substance. These techniques are more rapid and energy-efficient in the removal of water. Crowe et al. (1990) Cryobiol. 27:219-231. However, ambient temperature drying often yields denatured or even inactive substances unless an appropriate stabilizer is used. Crystallisation or co-precipitation can only be applied to a few substances, and the products of these methods have poor solubility. Additionally, there are problems in removing residual moisture.
Trehalose, α-D-glucopyranosyl- α-D-glucopyranoside, is a naturally occurring, inert, non-reducing and non-toxic disaccharide which was initially found to be associated with the prevention of desiccation damage in certain plants and animals which can dry out without damage and revive when rehydrated. Trehalose has been shown to be useful in preventing denaturation of a wide variety of substances such as proteins, viruses and foodstuffs during desiccation and subsequent storage. Formulations of products air dried in trehalose have been found to have a remarkably increased storage life. See U.S. Pat. Nos. 4,891,319; 5,149,653; 5,026,566; Blakely et al. (1990) Lancet 336:854; Roser (July 1991) Trends in Food Sci. and Tech., pp. 166-169; Colaco et al. (1992) Biotechnol. Internat., pp. 345-350; Roser (1991) BioPharm. 4:47; Colaco et al. (1992) Bio/Tech. 10:1007; Roser and Colaco (1993) New Scientist 138:25-28; and Crowe (1983) Cryobiol. 20:346-356. Trehalose also stabilizes lyophilized proteins, such as methanol dehydrogenase (Argall and Smith (1993) Biochem. Mol. Biol. Int. 30:491), and to confer thermoprotection to enzymes from yeast. Hottiger et al. (1994) Eur. J. Biochem. 219:187. Trehalose also inhibits the Maillard reaction between carbonyl groups of reducing sugars and amino groups of proteins. Loomis et al. (1979) J. Exp. Zool. 208:355-360; and Roser and Colaco (1993) New Scientist 138:24-28. Trehalose and a wide variety of stabilizing polyols have also been found useful in formulation of solid dosages.
There is a serious need for a method to inexpensively and stably incorporate substances into glass matrices with a minimum of residual moisture remaining in the product. Such a process would provide products exhibiting increased stability, a longer shelf life, and facile rehydration. Facile rehydration would be a particular advantage for parenterally administered pharmaceutical substances.
All references cited herein are hereby incorporated by reference.